ANCA-negative pauci-immune crescentic glomerulonephritis associated with nivolumab in lung cancer

Abstract

Immune checkpoint inhibitors (ICIs) have substantially improved the prognosis of many cancer patients but are also associated with various immune-related adverse events (irAEs). Kidney irAEs are relatively rare, with acute tubulointerstitial nephritis being the most common manifestation. However, some patients develop ICIs-associated glomerular diseases, including pauci-immune crescentic glomerulonephritis. In this report, we present the case of a 72-year-old man with lung squamous cell carcinoma treated with nivolumab, a monoclonal antibody targeting programmed cell death 1 (PD-1). The patient developed rapidly progressive glomerulonephritis a few weeks after initiating nivolumab therapy. Immunological tests yielded negative results, and a kidney biopsy revealed pauci-immune crescentic glomerulonephritis. Immunohistological examination confirmed programmed death ligand-1 (PD-L1) expression in the glomeruli. Despite intensive therapy, including corticosteroid pulse treatment, the patient’s kidney function did not recover, necessitating maintenance hemodialysis. This is the first report demonstrating PD-L1 staining in injured glomeruli caused by anti-PD-1 therapy. Immunohistochemistry for PD-L1 may aid in diagnosing glomerulonephritis related to anti-PD-1 therapy.

Introduction

Immune checkpoint inhibitors (ICIs) have substantially improved outcomes in various cancers and enhanced patient prognoses. In Japan, ICIs include monoclonal antibodies targeting programmed cell death protein 1 (PD-1; nivolumab and pembrolizumab), programmed cell death 1-ligand 1 (PD-L1; atezolizumab, durvalumab, and avelumab), and cytotoxic T-lymphocyte–associated antigen 4 (CTLA-4; ipilimumab and tremelimumab). Among these, nivolumab, the first approved ICI, is indicated for treating multiple cancers, including malignant melanoma, non-small cell lung cancer, renal cell carcinoma, classic Hodgkin’s lymphoma, head and neck cancer, gastric cancer, and malignant pleural mesothelioma [1]. ICIs exert antitumor effects by enhancing T-cell activity; however, they are also associated with immune-related adverse events (irAEs). Common irAEs for ICIs include hepatic dysfunction, interstitial pneumonia, colitis, neuropathy, and thyroid dysfunction. However, kidney-related irAEs are relatively rare. The incidence of kidney irAEs in ICI therapy is approximately 2.2%, with acute kidney injury (AKI) due to acute tubulointerstitial nephritis (ATIN) accounting for about 95% of these cases [2]. Recently, reports of ICI-associated glomerular diseases have been increasing, including pauci-immune glomerulonephritis (e.g., antineutrophil cytoplasmic antibody [ANCA]-associated vasculitis), anti-glomerular basement membrane (anti-GBM) disease, complement 3 (C3) glomerulonephritis, lupus nephritis, immunoglobulin (Ig) A (IgA) nephropathy, minimal change disease, amyloid A amyloidosis, and thrombotic microangiopathy [3]. To date, there have been no reports describing the deposition of PD-1/PD-L1 within the glomeruli in crescentic glomerulonephritis associated with PD-1/PD-L1 inhibitor therapy. This report describes a rare case of ANCA-negative, pauci-immune crescentic glomerulonephritis with PD-L1 staining in the injured glomeruli.

Case presentation

A 72-year-old man was diagnosed with squamous cell carcinoma of the left lower lobe 15 months before the onset of rapidly progressive glomerulonephritis (RPGN). His medical history included urolithiasis with persistent microscopic hematuria, though proteinuria had not been observed. The patient underwent a left lower lobectomy with lymph node resection via video-assisted thoracic surgery. Intraoperative pleural lavage cytology and the resection margin indicated positive results. He received four cycles of first-line chemotherapy with carboplatin and S-1. Three months after completing chemotherapy, computed tomography revealed left intrapulmonary, pleural, and lymph node metastases. Subsequently, nivolumab (240 mg) was administered 1 week before the onset of RPGN. At this time, his kidney function was normal (serum creatinine: 0.70 mg/dL), and urinalysis revealed no proteinuria but persistent isomorphic hematuria attributed to urolithiasis. One week after starting nivolumab therapy, his serum creatinine level increased to 0.92 mg/dL, and dipstick urinalysis revealed positive for proteinuria (3+). His serum creatinine levels progressively increased to 1.88 mg/dL and 5.15 mg/dL at 2 and 3 weeks post-nivolumab therapy, respectively. A 24-hour urine collection at 3 weeks revealed proteinuria (4.75 g/day). At this point, a nephrology consultation was sought. Immunological blood tests were negative for ANCAs, anti-GBM antibody, and antinuclear antibody and, with normal complement levels. Based on these findings, the patient was diagnosed with RPGN associated with an irAE, and was immediately hospitalized. A kidney biopsy was performed on the first day of hospitalization, and oral prednisolone (0.6 mg/kg/day, equivalent to 30 mg/day) was initiated the following day. However, due to oliguria (200–400 mL/day) and congestive heart failure, hemodialysis was initiated one day after starting steroid therapy.

Histological examination of the kidney revealed that 8 glomeruli exhibited global sclerosis (13%), and 25 glomeruli (42%) displayed cellular crescents out of a total of 60 glomeruli (Fig. 1A and B). Endocapillary proliferation and endothelial cell swelling were observed in some glomeruli (Fig. 1C). Moderate to severe tubular atrophy, interstitial fibrosis, and focal lymphocyte infiltration in the tubulointerstitial area were observed (Fig. 1A). Fibrinoid necrosis was not observed in the capillaries or small arteries of the kidney. Immunofluorescence examination revealed negative staining for immunoglobulins (IgG, IgA, and IgM) and complements (C3 and C1q) (Fig. 2). The electron microscopic study showed mostly normal findings, with rare electron-dense deposits (EDDs) observed in the paramesangial area (Fig. 3). Collectively, these findings were consistent with pauci-immune glomerulonephritis. Immunohistochemical staining for PD-L1 revealed focal PD-L1 expression in the crescentic glomeruli (predominantly in the mesangial cells) and tubulointerstitial lesions (Fig. 4A-C). Using the same PD-L1 staining protocol, we examined kidney tissues from a case of PD-1 inhibitor–associated tubulointerstitial nephritis (Fig. 4D–F) and a case of ANCA-associated crescentic glomerulonephritis (Fig. 4G–I). In both cases, no PD-L1 expression was observed within the glomeruli. (Fig. 4F and I) In the case of ANCA-associated crescentic glomerulonephritis, very weak and focal PD-L1 expression was detected in the cytoplasm of interstitial inflammatory cells. (Fig. 4H) In contrast, in the case of PD-1 inhibitor–associated tubulointerstitial nephritis, PD-L1 expression was clearly localized to the cell membranes of interstitial inflammatory cells. (Fig. 4E) Based on these findings, the prominent glomerular localization of PD-L1 observed in the present case appears to be a distinctive feature. As an additional investigation, we measured the serum concentrations of soluble PD-L1 and found a marked elevation to 197 pg/mL. On the 7th hospital day, the patient received intravenous pulse methylprednisolone (1000 mg daily for 3 days), followed by oral prednisolone at 0.8 mg/kg/day (equivalent to 40 mg/day). Despite intensive steroid therapies, severe proteinuria and hematuria persisted, and oliguria of approximately 200 mL per day continued. Consequently, his kidney function did not recover, and he required maintenance hemodialysis.

Fig. 1.

Kidney pathological findings. (A) Low-power magnification indicates tubular atrophy and focal inflammatory cell infiltration (Periodic acid-Schiff [PAS] stain, original magnification: 4×). (B) Representative image of a glomerulus with a cellular crescent (PAS stain, original magnification: 40×). (C) Some glomeruli display endocapillary proliferation, with capillary spaces narrowed by endothelial cell swelling and neutrophil proliferation (PAS stain, original magnification: 40×)

Fig. 2.

Immunofluorescence microscopic examination reveals negative staining for immunoglobulins and complements (original magnification: 40×)

Fig. 3.

Electron microscopy findings of the kidney biopsy. Podocyte foot process effacement and small electron-dense deposits (arrow) in the paramesangial area are observed (original magnification: 2500×)

Fig. 4.

Immunohistochemical staining for PD-L1. shows a low-magnification histological image (4×), while (B) and (C) are cropped high-magnification images (20×), with (C) stained using silver staining. Focal PD-L1 expression is observed in the injured glomeruli and interstitial lesions in (A) and (B). In the silver-stained image (C), PD-L1–positive areas (black arrows) are seen in the glomerular mesangial regions, corresponding to silver-positive areas. In contrast, PD-L1 expression is absent in the crescentic lesion in the lower right (gray arrow) and in podocytes in the upper left (gray arrow) (D) shows a low-magnification image (4×), and (E) and (F) are cropped high-magnification images (20×) of the interstitial lesion and glomerulus, respectively, from a case of PD-1 inhibitor–associated tubulointerstitial nephritis. (G) shows a low-magnification image (4×), and (H) and (I) are cropped high-magnification images (20×) of the interstitial lesion and glomerulus, respectively, from a case of crescentic glomerulonephritis secondary to ANCA-associated vasculitis. PD-L1 staining was performed using the following method: Paraffin-embedded 2-µm-thick kidney sections were stained with a monoclonal anti-PD-L1 antibody (Clone SP142, Abcam) using the Leica BOND-III staining platform. Heat antigen retrieval was performed using an antigen retrieval buffer (pH 9; BOND Epitope Retrieval Solution 2). After incubation with the primary antibody (dilution factor: 1:400), signals were visualized using the 3,3’-diaminobenzidine (DAB) system, as can be observed in this image

Due to a cytomegalovirus infection, the prednisolone dose was tapered, and he was discharged on the 56th hospital day (Fig. 5). Although nivolumab was not readministered, his kidney function remained impaired. The patient died of advanced lung cancer 10 months later.

Fig. 5.

Clinical course of this case. CBDCA, carboplatin; mPSL, methylprednisolone; PSL, prednisolone; UP, urinary protein; UCr, urinary creatinine

Discussion

This report describes ICIs-associated ANCA-negative pauci-immune crescentic glomerulonephritis, with confirmation of PD-L1 staining in the crescentic glomeruli. According to a previous systematic review of ICIs-associated glomerular diseases, pauci-immune glomerulonephritis/renal vasculitis (27%), minimal change disease/focal segmental glomerulosclerosis (24%), and C3 glomerulonephritis (5%) are among the most frequently reported types of glomerular diseases. The majority of ICIs-associated pauci-immune glomerulonephritis/vasculitis cases are characterized as focal segmental necrotizing glomerulonephritis or vasculitis. However, the pauci-immune crescentic glomerulonephritis presented in this case is exceedingly rare. Notably, most pauci-immune glomerulonephritis/renal vasculitis cases (80%) are serologically negative for ANCA [3]. One potential explanation is that autoantibodies induced by ICI administration may recognize epitopes different from typical autoantibodies, such as ANCA [4]. Among the reported 14 cases of crescentic glomerulonephritis associated with ICIs, 2 cases showed IgA nephropathy, 8 exhibited anti-GBM disease (including one case with concurrent membranous nephropathy), 3 showed pauci-immune crescentic glomerulonephritis with MPO-ANCA-associated vasculitis, and only 1 case was of ANCA-negative pauci-immune crescentic, similar to this case [4–17]. The “SUBARU-J Study,” which investigated real-world clinical outcomes of anticancer drug-related kidney injury in Japan using the J-RBR, was recently published [18]. Of the 135 cases, 46 were identified as PD-1 inhibitor–associated kidney injury. The majority of these cases (38 cases, 82.6%) were diagnosed as tubulointerstitial nephritis (TIN). In addition, there were 4 cases of IgA nephropathy, 2 cases of minimal change disease, 1 case of thrombotic microangiopathy (TMA), and 1 case of ANCA-negative crescentic glomerulonephritis (CrGN), which corresponds to the present case.

Previous studies have reported a latency period of 2 to 10 months before the onset of ICIs-associated AKI caused by ATIN [19, 20]. In this case, glomerulonephritis developed in less than a week after nivolumab administration, which is shorter than the reported latency period. This raises the possibility that crescentic glomerulonephritis might be associated with the patient’s underlying lung cancer rather than solely caused by nivolumab. Indeed, ANCA-positive pauci-immune crescentic glomerulonephritis/vasculitis is relatively common in association with malignancies [21]. However, cases of ANCA-negative pauci-immune crescentic glomerulonephritis associated with malignancy are rare.

In previous reports, PD-L1 immunohistochemical staining has been used to examine the association between PD-1 and crescentic glomerulonephritis. Cassol et al. reported that PD-L1 could be detected in renal tubular cells and suggested its utility in diagnosing patients with PD-1 inhibitor-related ATIN [22]. Conversely, another study suggests that PD-L1 expression levels in the kidney vary significantly among individuals, even under normal conditions, and that high levels of PD-L1 expression may be associated with the development of kidney irAEs following anti-PD-1 antibody therapy [23]. In this case, PD-L1 was expressed in tubulointerstitial lesions with severe inflammatory cell infiltration and in nearly all glomeruli with crescents. Multiplex immunofluorescence staining was performed, but it was not possible to clearly assess which cells expressed PD-L1. Generally, PD-L1 is expressed in some normal cells involved in the immune system, aside from cancer or inflammatory cells. In normal kidneys, PD-L1 is expressed in proximal tubular epithelial cells, interstitial dendritic cells, and macrophages. However, PD-L1 expression in the glomeruli, including podocytes, remains unclear [24]. PD-L1 is essentially an immunosuppressive protein that resides in the cell membrane and binds to PD-1 on T cells. However, recent studies have shown that PD-L1 is also present in the cytoplasm and is now recognized to be internalized from the membrane into the cell via endocytosis [25]. Although the precise role of intracellular PD-L1 remains unclear, accumulating evidence suggests a potential link to therapeutic efficacy and resistance. In this regard, a study of 31 patients with malignant melanoma reported that those who developed severe irAEs during anti–PD-1 antibody therapy exhibited a significant increase and greater variability in serum sPD-L1 levels compared with those who did not [26]. While endocytosis of soluble PD-L1 has not yet been reported, it is conceivable that elevated circulating sPD-L1 could be internalized into glomerular cells and contribute to glomerular injury. Given the markedly high serum sPD-L1 level in the present case, it is plausible that such a mechanism may have been involved in the development of severe crescentic glomerulonephritis. Further experimental studies are warranted to clarify the mechanisms by which soluble PD-L1 is endocytosed and its role in the pathogenesis of crescentic glomerulonephritis.

When an irAE is suspected, it is recommended to discontinue ICIs immediately and initiate steroid therapy, depending on the severity of the irAE [27]. However, this recommendation primarily applies to ICIs-associated ATIN, and standardized treatment options for ICIs-associated glomerulonephritis remain unclear. In the 14 previously reported cases of ICIs-associated crescentic glomerulonephritis, ICIs were discontinued and steroid therapy was administered in all cases. Complete recovery of kidney function was observed in only one case, while eight cases showed partial recovery, and five cases exhibited no improvement in kidney function [4–17]. Among these cases, a 71-year-old woman with lung adenocarcinoma developed nephrotic-range proteinuria and microscopic hematuria with stable kidney function (serum creatinine: 0.8 mg/dL) after 10 months of pembrolizumab (anti-PD-1 antibody) therapy. The immunological test and kidney biopsy results indicated ANCA-negative pauci-immune crescentic glomerulonephritis. Her hematuria and proteinuria improved following high-dose steroid treatment [10].

Although immunofluorescence did not reveal clear deposition of immunoglobulins or complement components in the glomeruli, electron microscopy demonstrated a small amount of EDDs in the paramesangial area. In pauci-immune crescentic glomerulonephritis, particularly in ANCA-associated cases, EDDs have been reported in approximately 54% of renal biopsies. Moreover, the presence of EDDs has been associated with higher serum creatinine levels, greater 24-hour urinary protein excretion, and a higher percentage of glomeruli with crescents, suggesting a correlation with poorer renal outcomes [28]. In the present case, the presence of paramesangial EDDs may have contributed to the progression to end-stage renal disease despite intensive steroid therapy.

This study has certain limitations. There are three enzymatic methods for detecting ANCA, including enzyme-linked immunosorbent assay (ELISA), chemiluminescent enzyme immunoassay (CLEIA), and fluoroenzyme immunoassay (FEIA). In this case, ANCAs were not detected using CLEIA; however, different results may have been obtained with other assays, such as ELISA or FEIA. Moreover, considering that most cases of ICIs-associated pauci-immune glomerulonephritis/vasculitis are ANCA-negative, these patients may possess different types of ANCAs or novel autoantibodies beyond MPO- or PR3-ANCAs [29]. Minor ANCAs, such as BPI-ANCA and elastase-ANCA, were not measured in this case, and their involvement cannot be ruled out.

In conclusion, we reported a case of ANCA-negative pauci-immune crescentic glomerulonephritis associated with nivolumab, where PD-L1 expression in the glomeruli was demonstrated by immunostaining and may be associated with crescent formation. Since crescentic glomerulonephritis as an irAE is rare and its mechanism is not well understood, further case series are necessary to elucidate this condition.

Author contributions

R.F., M.E., and K.T. conceived and designed research; M.K., Y.B., H.T., T.U., H.T., M.N., and T.K. performed treatment; R.F. and K.S. performed histological diagnosis; R.F. and M.E. drafted manuscript; H.T., T.U., H.T., F.F., M.N., T.K., K.T., M.M., and K.T. edited and revised manuscript.

Declarations

Conflict of interest

All the authors have declared no competing interest.

Ethical approval

All procedures in this study involving human participants were performed in accordance with the ethical standards of the institutional review board and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards.

Informed consent

Informed consent was obtained from all individual participants included in the case report.